Open AccessCBS mutations are good predictors for B6‐responsiveness: A study based on the analysis of 35 Brazilian Classical Homocystinuria patients
Author(s) -
Poloni Soraia,
SperbLudwig Fernanda,
Borsatto Taciane,
Weber Hoss Giovana,
Doriqui Maria Juliana R.,
Embiruçu Emília K.,
BoaSorte Ney,
Marques Charles,
Kim Chong A.,
Fischinger Moura de Souza Carolina,
Rocha Helio,
Ribeiro Marcia,
Steiner Carlos E.,
Moreno Carolina A.,
Bernardi Pricila,
Valadares Eugenia,
Artigalas Osvaldo,
Carvalho Gerson,
Wanderley Hector Y. C.,
Kugele Johanna,
Walter Melanie,
GallegoVillar Lorena,
Blom Henk J.,
Schwartz Ida Vanessa D.
Publication year - 2018
Publication title -
molecular genetics and genomic medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.765
H-Index - 29
ISSN - 2324-9269
DOI - 10.1002/mgg3.342
Subject(s) - homocystinuria , cystathionine beta synthase , missense mutation , exon , genetics , microbiology and biotechnology , gene , genotype , intron , mutation testing , mutation , medicine , point mutation , biology , amino acid , methionine
Background Classical homocystinuria ( HCU ) is a monogenic disease caused by the deficient activity of cystathionine β‐synthase (CβS). The objective of this study was to identify the CBS mutations in Brazilian patients with HCU . Methods gDNA samples were obtained for 35 patients (30 families) with biochemically confirmed diagnosis of HCU . All exons and exon‐intron boundaries of CBS gene were sequenced. Gene expression analysis by qRT ‐ PCR was performed in six patients. Novel missense point mutations were expressed in E. coli by site‐directed mutagenesis. Results Parental consanguinity was reported in 16 families, and pyridoxine responsiveness in five (15%) patients. Among individuals from the same family, all presented the same phenotype. Both pathogenic mutations were identified in 29/30 patients. Twenty‐one different mutations were detected in nine exons and three introns; being six common mutations. Most prevalent were p.Ile278Thr (18.2%), p.Trp323Ter (11.3%), p.Thr191Met (11.3%), and c.828+1G>A (11.3%). Eight novel mutations were found [c.2T>C, c.209+1delG, c.284T>C, c.329A>T, c.444delG, c.864_868del GAG c.989_991del AGG , and c.1223+5G>T]. Enzyme activity in E. coli‐ expressed mutations was 1.5% for c.329A>T and 17.5% for c.284T>C. qRT ‐ PCR analysis revealed reduced gene expression in all evaluated genotypes: [c.209+1delG; c.572C>T]; [c.2T>C; c.828+1G>A]; [c.828+1G>A; c.1126G>A]; [c.833T>C; c.989_991del AGG ]; [c.1058C>T; c.146C>T]; and [c.444delG; c.444delG]. The expected phenotype according to the genotype (pyridoxine responsiveness) matched in all cases. Conclusions Most patients studied were pyridoxine nonresponsive and presented early manifestations, suggesting severe phenotypes. Many private mutations were observed, but the four most prevalent mutations together accounted for over 50% of mutated alleles. A good genotype–phenotype relationship was observed within families and for the four most common mutations.